Increasingγ-CD conversion rates by improving thermostability of Bacillus sp.FJAT-44876γ-CGTase  

作　　者：Xiaoxiao Li Danni Zheng Jing Wu Zhengyu Jin Birte Svensson Yuxiang Bai 

机构地区：[1]State Key Laboratory of Food Science and Technology,Jiangnan University,Wuxi,Jiangsu,214122,China [2]School of Food Science and Technology,Jiangnan University,Wuxi,Jiangsu,214122,China [3]Collaborative Innovation Center of Food Safety and Quality Control in Jiangsu Province,Jiangnan University,Wuxi,Jiangsu,214122,China [4]Enzyme and Protein Chemistry,Department of Biotechnology and Biomedicine,Technical University of Denmark,DK-2800 Kgs,Lyngby,Denmark

出　　处：《Food Bioscience》2023年第1期346-354,共9页食品生物科学（英文）

基　　金：supported by National Natural Science Foundation of China(No.32072268 and No.32201967);Natural Science Foundation of Jiangsu Province(BK20211581 and BK20221072);the International Joint Research Laboratory for Starch Related Enzymes at Jiangnan University with DTU Bioengineering。

摘　　要：Poor thermostability is a limiting factor for applications ofγ-CGTase that affects starch utilization and yield ofγ-CD.Here thermostability of Bacillus sp.FJAT-44876γ-CGTase(BFγ-CGTase)was improved by addition of Ca^(2+)and site-directed mutagenesis.Thus,10 mM Ca^(2+)increased the half-life(t1/2)at 55 and 60℃ from 3.0 to 0.3 h to 17.4 and 2.0 h,respectively.Fluorescence spectra indicated that Ca^(2+)stabilized the tertiary structure of the BFγ-CGTase and hence improved the thermostability.Mutation to serine of a glycine residue in anα-helix related to thermostability also improved the stability especially at the higher temperature.Importantly,in 10 mM Ca^(2+)the G208S mutant further increased t1/2 to 20.0 h and 4.0 h at 55 and 60℃,respectively.The G208S mutant in 10 mM Ca^(2+)producedγ-CD from tapioca starch with 40%increased yield of that from BFγ-CGTase.This work involving rational protein engineering provided a new tool for enzymaticγ-CD production.

关 键 词：STARCH Protein engineering Three-dimensional structure 

分 类 号：TS201.3[轻工技术与工程—食品科学]